大黄鱼过氧化物酶Ⅳ在人胚肾细胞中的表达与抗氧化活性测定

为了研究大黄鱼（Pseudosciana crocea）peroxiredoxin Ⅳ（Lyc-Prx Ⅳ）在细胞内的抗氧化功能,构建了表达大黄鱼Prx Ⅳ的重组质粒p CMV-Lyc-Prx Ⅳ,瞬时转染人胚肾细胞（HEK-293T）,利用Western blotting方法检测转染后的细胞样品中Lyc-Prx Ⅳ的表达情况,并通过测定细胞内过氧化氢浓度来评价Lyc-Prx IV的体内抗氧化作用.结果显示,Lyc-Prx Ⅳ可在转染重组质粒p CMV-Lyc-Prx Ⅳ的HEK-293T细胞中表达,且细胞中A560处的吸光值在转染后6、12、24 h后分别为0.154、0.116以及0.162,而瞬时转染空载体p CMV后细胞样品在A560处的吸光值在转染后一直稳定在0.260左右,说明细胞中过氧化氢的浓度在转染后6、12、24 h时明显低于瞬时转染空载体p CMV细胞中的过氧化氢浓度（p〈0.01）.表明Lyc-Prx Ⅳ在生物体内可以分解过氧化氢,参与生物体内氧化还原状况的调控.     

大菱鲆两种转基因方法的比较研究

对外源基因导入大菱鲆(Scophthat musmaximus)的两种方法进行了研究,采用经过改进的电脉冲方法以及哺乳动物细胞中常用的转染法导入外源基因,并对两种方法进行比较。电脉冲最佳导入条件为:脉冲电压为300V/cm,脉冲次数为5,外源DNA质量浓度为20mg/L;转染法的最佳操作时间为受精后50min。应用上述两种方法进行批量转基因鱼实验,得到外源基因的导入率分别为20%～28%和60%～70%。实验表明利用转染试剂jetPEI处理单细胞期的大菱鲆受精卵能得到较高的孵化率及较高的转基因效率。     

紫球藻溶气气浮法采收条件研究

探讨溶气气浮法采收紫球藻细胞的可行性,较为详细地研究了气浮操作参数对气浮采收效果的影响,确定合适的紫球藻细胞溶气气浮采收工艺条件。结果表明,通过调节pH值,能够获得藻细胞絮体,从而为无絮凝剂添加的紫球藻细胞气浮采收创造了条件。确定的最佳溶气气浮工艺为:pH值10.1,溶气压力0.6MPa,溶气时间17h,溶气水进水流量60L/h,溶气水/原料藻液体积比1.0。在此条件下,紫球藻的采收效果很好,细胞采收率可达到90%以上,浓缩倍数可达到4左右。培养液藻细胞浓度的增加对细胞采收率影响不大,但会降低浓缩倍数。     

气升式反应器培养集胞藻6803过程中光能利用特性研究

在气升式光生物反应器中,对集胞藻 6803 分批培养条件下的光衰减、平均光强、光的分布以及藻细胞生长对光能的利用特性进行分析.结果表明:Lambert-Beer 定律可较好描述细胞浓度及光程对光衰减的影响;随着藻细胞密度的增加,光生物反应器内平均光强不断减小,藻细胞的比光能吸收速率和比生长速率也不断降低,反应器中"暗区"体积在培养 6 d 后超过总体积的 80%;利用 Pirt 细胞生长生物能量模型描述比生长速率和比光能吸收速率的关系,在入射光强为 58.73 KJ/(m2(h),求得基于光能的细胞得率 YG 和维持系数 m 分别为 9.98×10-3g/kJ和0.445kJ/(g(h).     

五种中草药对海洋光合细菌生长影响的研究

研究了不同方式提取的中草药浸提液以及不同添加量的浸提液对海洋先合知菌生长的影响。结果表明对甘草、黄芩、栝楼等中草药水煮法效果较好,而对青蒿、党参等中草药醇提法效果较好,同时发现上述几种中草药对海洋光合细菌的生长均有促进作用。其中添加4.0%,4.0%和1.0%的栝楼、黄苓和党参浸提液的细菌经过120h的培养.细菌数分别比对照组提高了99.1%、68.8%和68.4%,甘草,青蒿.比对照组提高了10.9%和6.9%,其最佳添加量均为0.5%。     

海鞘血细胞培养和细菌污染控制

以海鞘(Ciona savignyi)为研究对象,考察了海鞘血细胞原代培养方法及培养中细菌污染的鉴定和控制.血细胞在培养1 h后贴壁,细胞基本以圆形细胞为主,变形细胞一般呈不规则形.变形细胞存活时间较短,而圆形细胞存活时间较长.对于培养过程中的细菌污染,通过细菌分离、培养和纯培养发现两类菌株检出率较高,均为革兰氏阴性菌.经PCR扩增16S rDNA基因序列片断,结果显示这两类菌株分别属于弧菌属(Vibrio)和施万氏菌属(Shewanella).药敏试验结果显示,弧菌属对氯霉素和环丙沙星等具有较强的敏感性;对施万氏菌属较敏感的药物依次为亚胺培南和氯霉素等.最后比较了几种抗生素组合控制血细胞培养中细菌污染的使用效果,其中氯霉素、亚胺培南加双抗的组合有较好的抑菌效果并对培养细胞的贴壁和生长没有影响.     

抑酸复合茶对高尿酸血症模型大鼠肾功能的影响及降尿酸作用机制研究

目的：采用高尿酸血症模型大鼠研究抑酸复合茶对高尿酸血症的防治作用,并探讨其降尿酸的作用机制。方法：大鼠适应性饲养1周后根据随机分组原则分为正常对照组、模型组、别嘌醇组及抑酸复合茶高、低剂量组,每组各12只。除正常对照组外,其余各组均采用腺嘌呤联合氧嗪酸钾连续灌胃3周以建立高尿酸血症模型,建模同时给予药物干预,观察各组大鼠给药前后血尿酸（UA）及给药后尿素氮（BUN）、肌酐（Scr）、胱抑素C（CysC）、一氧化氮（NO）的变化。采用Western blot法检测抑酸复合茶对HEK293细胞URAT1和GLUT9表达的影响。结果：给药后,与模型组比较,抑酸复合茶高剂量组UA、BUN、Scr、CysC均降低,NO升高,差异均有统计学意义（P<0.05）;低剂量组UA、BUN降低,NO升高,差异亦有统计学意义（P<0.05）;与别嘌醇组比较,抑酸复合茶高剂量组CysC下降,NO升高,差异有统计学意义（P<0.05）;抑酸复合茶还可使HEK293细胞中URAT1和GLUT9的表达量降低。结论：抑酸复合茶对高尿酸血症大鼠降尿酸作用明显,且对血管内皮细胞和肾功能具有一定的保护作用;其作用机制可能是通过下调肾脏URAT1和GLUT9蛋白表达而发挥。     

大菱鲆红体病虹彩病毒体外增殖条件的研究

探讨大菱鲆病毒疫苗的研制,以牙鲆鳃细胞(FG细胞)为增殖体系,利用细胞病变效应(CPE)为判断指标,对大菱鲆红体病虹彩病毒(Turbot reddish body iridovirus,TRBIV)的体外最适增殖条件等进行了研究。结果表明,病毒接种量、病毒吸附时间、细胞培养条件对TRBIV的体外增殖均有明显的影响,以0.2 mL病毒液接种FG细胞、吸附2 h、用含10%胎牛血清的MEM培养液于22℃培养时,病毒的增殖速度最快。研究还发现,病毒的收获时间和病毒液冻融次数对所得病毒的感染力均有显著影响,在最佳增殖条件下,病毒在FG细胞中增殖至第4天时进行收获,且只冻融1次,其感染活性最强,反复冻融后病毒的感染活性会显著降低。     

Δ6-去饱和酶和C20-延伸酶抑制乳腺癌细胞MCF-7增殖的研究

为探讨外源基因Δ6-去饱和酶与C20-延伸酶基因对肿瘤细胞增殖的抑制作用,本研究选取了微拟球藻(Nannochloropsis oculata)作为研究对象,对其Δ6-去饱和酶与C20-延伸酶基因进行了逆转录PCR扩增,并将获得的上述2个基因的编码区与真核表达质粒pEGFP-C3构建重组表达质粒,转染乳腺癌细胞MCF-7,以pEGFP-C3为对照,培养48h后,在荧光显微镜下观察转染细胞中的GFP荧光信号以判断重组质粒表达情况,荧光实时定量PCR分析转染细胞中外源基因的转录水平,MTT比色法检测外源基因对MCF-7细胞的增殖抑制程度,高效气相色谱检测转染细胞中多不饱和脂肪酸(Polyunsaturated fatty acids,PUFAs)的含量变化。结果表明2个外源基因均可在MCF-7细胞中表达,并能有效地抑制MCF-7细胞的增殖,改变MCF-7细胞中2种PUFAs的相对含量,且n-3PUFAs与n-6PUFAs的比值升高。推测过表达的Δ6-去饱和酶与C20-延伸酶可能通过改变乳腺癌细胞MCF-7细胞膜中PUFAs的相对含量,达到抑制乳腺癌细胞MCF-7增殖的效果。     

紫菜细胞悬浮培养

用酶解技术获得条斑紫菜离体细胞并对其进行细胞悬浮培养研究;对藻休不同部位的细胞分化发育方式,不同悬浮器具培养效果及不同起始培养的细胞密度进行了探索。结果表明:用气升式生物反应器培养细胞效果最佳,起始培养的细胞密度可达l×10~5细胞/ml,细胞存活率达85%。     

Microbial Biomass and Organic Nutrients in the Deep-Sea Sediments of the Central Indian Ocean Basin

In order to assess the impact of deep-sea mining on the in situ benthic life, we measured the microbial standing stock and concentration of organic nutrients in the deep-sea sediments of the Central Indian Ocean Basin in the Indian pioneer area. Sediments were collected using box core and grab samples during September 1996. The total bacterial numbers ranged from 10 10 -10 11 cells per g -1 dry weight sediment. There was a marginal decrease in the number of bacteria from surface to 30 cm depth, though the subsurface section registered a higher number than did the surface. The highest numbers were encountered at depths of 4-8 cm. The retrievable number of bacteria were two orders less in comparison with the direct total counts of bacteria. An almost homogeneous distribution of bacteria, total organic carbon, living biomass, and lipids throughout the depth of cores indicates active microbial and benthic processes in the deep sea sediments. On the other hand, a uniform distribution of total counts of bacteria, carbohydrates, and total organic carbon in all the cores indicates their stable nature and suggests that they can serve as useful parameters for long-term monitoring of the area after the benthic disturbance. Further studies on temporal variability in this region would not only verify the observed norms of distribution of these variables but would also help to understand restabilization processes after the simulated benthic disturbance.     

Assessing the Validity of Seismo-Acoustic Predictor Equations for Obtaining Seabed and Sub-Surface Sediment Physical Properties

The interdependence between the seismo-acoustic properties of a marine sediment and its geotechnical/physical parameters has been known for many years, and it has been postulated that this should allow the extraction of geotechnical information from seismic data. Though in the literature many correlations have been published for the surficial layer, there is a lack of information for greater sediment depths. In this article, a desktop study on a synthetic seafloor model illustrates how the application of published near-surface prediction equations to subsurface sediments (up to several tens of meters burial depth) can lead to spurious predictions. To test this further, acoustic and geotechnical properties were measured on a number of sediment core samples, some of which were subjected to loading in acoustically-equipped consolidation cells (oedometers) to simulate greater burial depth conditions. For low effective pressures (representing small burial depths extending to around 10 meters subsurface), the general applicability of established relationships was confirmed: the prediction of porosity, bulk density, and mean grain size from acoustic velocity and impedance appears generally possible for the investigated sedimentary environments. As effective pressure increases through, the observed relationships deviate more and more from the established ones for the near-surface area. For the samples tested in this study, in some instances increasing pressure even resulted in decreasing velocities. There are several possible explanations for this abnormal behavior, including the presence of gas, overconsolidation, or bimodal grain size distribution. The results indicate that an appropriate depth correction must be introduced into the published prediction equations in order to obtain reliable estimates of physical sediment properties for greater subsurface depths.     

A Review of Some Environmental Issues Affecting Marine Mining

This article reviews information recently available from existing marine and coastal mining for responses to environmental issues affecting marine mining at different depths. It is particularly but not exclusively concerned with those issues affecting seabed biodiversity impact and recovery. Much information has been gathered in the past 10 years from shallow mining operations for construction aggregate, diamonds, and gold, from coastal mines discharging tailings to shallow and deep water, and from experimental deep mining tests. The responses to issues identified are summarized in a series of eight tables intended to facilitate site-specific consideration. Since impacts can spread widely in the surface mixing layer SML, and can affect the biologically productive euphotic zone, the main issues considered arise from the depth of mining relative to the SML of the sea. Where mining is below the SML, the issue is whether it is environmentally better to bring the extraction products to the surface vessel for processing (and waste discharge), or to process the extraction products as much as possible on the seabed. Responses to the issues need to be sitespecific, and dependent on adequate preoperational environmental impact and recovery prediction. For deep tailings disposal from a surface vessel, there are four important environmental unknowns: (1) the possible growth of "marine snow" (bacterial flocs) utilizing the enormous quantities of fine tailings particles (hundreds or thousands of metric tons per day) as nuclei for growth, (2) the possibility that local keystone plankton and nekton species may migrate diurnally down to and beyond the depth of deep discharge and hence be subjected to tailings impact at depth, (3) the burrow-up capability of deep benthos and their ability to survive high rates of tailings deposition, and (4) the pattern and rate of dispersion of a tailings density current through the deep water column from discharge point to seabed. Actions to obtain relevant information in general and site-specifically are suggested.     

Increased Particle Fluxes at the INDEX Site Attributable to Simulated Benthic Disturbance

Particle fluxes were measured 7 m above the sea bottom during the predisturbance, disturbance, and postdisturbance periods by using time series sediment traps attached to seven deep-sea moorings deployed in the INDEX experiment site in the Central Indian Basin. The predisturbance particle fluxes varied between 22.3 to 55.1 mg m -2 day -1 . Increased and variable particle fluxes were recorded by the sediment traps during the disturbance period. The increase observed was 0.5 to 4 times more than the background predisturbance fluxes. The increases in particle fluxes (~4 times) recorded by the sediment trap located in the southwestern direction (DMS-1) were the greatest, which could be the result of preferential movement of resuspended particles generated during the deep-sea benthic disturbance along the general current direction prevailing in this area during the experimental period. Also, the traps located closer to the disturbance area recorded greater fluxes than did the traps far away, across the Deep Sea Sediment Resuspension System path. This variability in recorded particle fluxes by the traps around the disturbance area clearly indicates that physical characteristics such as grain size and density of the resuspended particles produced during the disturbance had an important effect on particle movement. The postdisturbance measurements during ~5 days showed a reduction in particle fluxes of ~50%, indicating rapid particle settlement.     

Geoacoustic Inversion via Genetic Algorithm and Its Application to Manganese Sediment Identification

An acoustic inversion method using a wide-band signal and two near field receivers is proposed and applied to multiple layered seabed models including a manganese sediment. The inversion problem can be formulated into a probabilistic model comprised of signals, a forward model, and additive noise. The forward model simulates wide-band signals, such as chirp signals, and is chosen to be the source-waveletconvolution plane wave modeling method. The wavelet matching technique, using weighted least-squares fitting, estimates the sediment sound-speed and thickness on which determination of the possible numerical ranges for a priori uniform distribution is based. The genetic algorithm is applied to a global optimization problem to find a maximum a posteriori solution for determined a priori search space. Here the object function is defined by an L 2 norm of the difference between measured and modeled signals. Not only the marginal pdf but also its statistics are calculated by numerical evaluation of integrals using the samples selected during importance sampling process of the genetic algorithm.     

海上大直径钢管桩打桩过程中桩周土强度弱化研究

海上大直径钢管桩打桩过程中,桩周土体受到强烈扰动而发生强度弱化,掌握桩周土体强度弱化规律对于准确预测打桩过程、保证工程安全具有重要意义。为研究土体强度弱化规律,开展了环剪试验模拟打桩对桩周土体的扰动,测试土体强度随剪切速率的变化规律,建立了描述土体强度弱化规律的拟合公式,引入到打桩分析软件中。研究结果表明：土体的强度折减程度不仅与土体本身的性质有关还受到土体的埋深和剪切速率的影响,埋深越深土体强度折减程度越低,剪切速率越高土体强度折减越高,在打桩分析中可采用这里推荐的线性折减方法来模拟不同深度处土体强度的折减规律。     

Determination of In Situ Sediment Shear Strength from Advanced Piston Corer Pullout Forces

The advanced piston corer (APC) has been used by the Ocean Drilling Program since 1985 for recovering soft sediments from the ocean floor. The pullout force measured on extracting the core barrel from the sediment is shown to correlate with the average shear strength of the sediment core measured in the ship's laboratory. A simple rule of thumb is derived relating the shear strength of the sediment to the pullout force. Multiple APC holes at individual sites allow the consistency of the pullout measurements to be assessed. The effects of different operational procedures during APC coring are also explored. Although generally applicable, the correlation between pullout force and laboratory measurements of shear strength breaks down for some APC holes, possibly because of the disturbance of some sediment types during the APC coring process. A better understanding of the physical process of APC coring, and its effect on the properties of the sediment both inside and immediately outside the core barrel, would indicate what confidence can be put on the measurement of pullout force as a way of evaluating the in situ shear strength of deep sea sediments.     

Interannual Sea Level Variations and Annual Tides in the Northwestern Pacific

Specific properties of the interannual sea level variations and annual tides in the Northwestern Pacific were studied. Several tide stations were monitored. The monthly mean sea level for the year of 1995 was analyzed at each tide station. A seismic event in 1995, some tectonic activity around the subject area, and the Kuroshio (the oceanic western boundary current) may possibly contaminate results which would have occurred from the astronomical annual tide alone.     

Flow Cytometric Analysis of Coastal Lagoon Bacterioplankton and Picophytoplankton: Fixation and Storage Effects

The effect of different fixation and storage protocols on the flow cytometric (FCM) simultaneous analysis of bacterioplankton and phytoplankton in coastal seawater samples (Mediterranean coastal lagoons) was investigated. FCM measurements (cell number, fluorescence and scatter characteristics) were obtained through DAPI staining. Three fixatives [glutaraldehyde (GA), formaldehyde (FA) and paraformaldehyde (PFA)] and two storage (3 months duration) methods (5 °C and −196 °C) were tested. Two dominant populations were detected in studied samples: bacteria and eukaryotic picophytoplankton. Adding fixatives (2% final concentration) appears necessary to obtain FCM exhaustive counts of all the bacteria and phytoplanktonic cells. This was related to the permeation effect of fixatives which allowed a better DAPI staining of the cells. Maximum fluorescence, i.e. optimal staining of the cells was obtained with FA or PFA, and significant lower fluorescences with GA. Fixed samples stored at 5 °C induced rapid cell loss. Only storage in liquid nitrogen of samples fixed with FA or PFA, allows mid-term (≥4 months) preservation of bacteria or picophytoplankton cell numbers, and limited evolution of DAPI-induced fluorescence and scatter characteristics.